title: Extracellular Ca²+ alleviates NaCl-induced stomatal opening through a pathway involving H₂O₂-blocked Na+ influx in Vicia guard cells.
abstract: To gain further insights into the function of extracellular Ca²+ in alleviating salt stress, Vicia faba guard cell protoplasts (GCPs) were patch-clamped in a whole-cell configuration. The results showed that 100 mM NaCl clearly induced Na+ influx across the plasma membrane in GCPs and promoted stomatal opening. Extracellular Ca²+ at 10 mM efficiently blocked Na+ influx and inhibited stomatal opening, which was partially abolished by La³+ (an inhibitor of plasma membrane Ca²+ channel) or catalase (CAT, a H₂O₂ scavenger), respectively. These results suggest that the plasma membrane Ca²+ channels and H₂O₂ possibly mediate extracellular Ca²+-blocked Na+ influx in GCPs. Furthermore, extracellular Ca²+ activated the plasma membrane Ca²+ channels under NaCl stress, which was partially abolished by CAT. These results, taken together, indicate that hydrogen peroxide (H₂O₂) likely regulates Na+ uptake by activating plasma membrane Ca²+ channels in GCPs. In accordance with this hypothesis, H₂O₂ could mimic extracellular Ca²+ to activate Ca²+ channels and block Na+ influx in guard cells. A single-cell analysis of cytosolic free Ca²+ ([Ca²+](cyt)) using Fluo 3-AM revealed that extracellular Ca²+ induced the accumulation of cytosolic Ca²+ under NaCl stress, but had few effects on the accumulation of cytosolic Ca²+ under non-NaCl conditions. All of these results, together with our previous studies showing that extracellular Ca²+ induced the generation of H₂O₂ in GCPs during NaCl stress, indicate that extracellular Ca²+ alleviates salt stress, likely by activating the H₂O₂-dependent plasma membrane Ca²+ channels, and the increase in cytosolic Ca²+ appears to block Na+ influx across the plasma membrane in Vicia guard cells, leading to stomatal closure and reduction of water loss.
authors.affiliation: Key Laboratory of Plant Stress Biology, College of Life Sciences, Henan University, 85 Minglun Street, Kaifeng 475004, People's Republic of China.
authors.forename: Xiang
authors.initials: X
authors.lastname: Zhao
authors.forename: Ya-Jing
authors.initials: YJ
authors.lastname: Wang
authors.forename: Yan-Liang
authors.initials: YL
authors.lastname: Wang
authors.forename: Xi-Li
authors.initials: XL
authors.lastname: Wang
authors.forename: Xiao
authors.initials: X
authors.lastname: Zhang
mesh_terms: D001692:Biological Transport; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology; D006861:Hydrogen Peroxide / Q000378:metabolism*; D015640:Ion Channel Gating; D054046:Plant Stomata / Q000378:metabolism* / Q000502:physiology; D011523:Protoplasts / Q000378:metabolism; D012964:Sodium / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology; D013312:Stress, Physiological; D031307:Vicia faba / Q000378:metabolism*
chemical_list: D015220:Calcium Channels; D012965:Sodium Chloride; D012964:Sodium; D006861:Hydrogen Peroxide; D002118:Calcium